PROJECT SUMMARY We study how injury and inflammation induce mature cells like the digestive-enzyme-secreting zymogenic chief cell (ZC) to disassemble their complex cell architecture and re-enter the cell cycle. We previously showed that ZCs become proliferative via a sequence of molecular-cellular events conserved across many tissues and species in scenarios where mature cells are recruited back into the cell cycle in response to tissue damage. Thus, cells have an evolutionarily conserved program for this reprogramming, as they do for death (apoptosis) and division (mitosis). We call this program paligenosis and showed that mature cells: first degrade/recycle their differentiated cell components (Stage 1), then induce expression of progenitor-like genes (eg. Sox9 = Stage 2), and finally re-enter the cell cycle (Stage 3). Paligenotic ZCs convert to cells that can be seen histopathologically as the type of metaplasia that occurs in stomach during long-term infection with the bacterium Helicobacter pylori: pseudopyloric or Spasmolytic Polypeptide Expressing Metaplasia (SPEM). Metaplasia can either resolve as tissue is repaired or become chronic and increase risk for progression to dysplasia and cancer. We have shown that paligenosis is governed by dynamic changes in mTORC1, the cellular translation control protein complex. mTORC1 is elevated at baseline in ZCs to drive translation of digestive enzymes, it shuts off at Stage 1, and reactivates at Stage 3. Without mTORC1, paligenosis stops at Stage 2 with cells looking metaplastic, but unable to enter S-phase. Here, we explore the mechanisms that induce and promote paligenosis. We show preliminary data implicating the Integrated Stress Response (ISR) pathway as a central paligenosis hub with a particular role for the transcription factor Atf3, which is associated with the ISR, and another gene which we hypothesize is a target of ATF3: Ifrd1, a multifunctional scaffolding protein. We hypothesize that the stress of large-scale tissue damage and/or inflammation triggers ISR hyperactivity, which leads to greatly increased ATF3 and IFRD1, to help push cells back into the cell cycle. In the absence of Atf3 or Ifrd1, we show paligenosis is defective. Our Specific Aims will be: 1) to confirm and further characterize at which stages ISR is active and confirm and characterize the role for ATF3 using, in part Atf3?/? mice; 2) to identify additional genes involved in the ISR and paligenosis by confirming the role of IFRD1 with Ifrd1?/? mice, probe relative contributions of ATF3 and IFRD1 by characterizing double knockout mice, and finally to perform ChIP- and RNA- Seq during paligenosis ATF3; 3) to test known ATF3 and ISR genes and new targets developed in Aim 2 in a pipeline of more physiological disease models (eg chronic infection of mice with H pylori), human translational samples (Tissue Microarray and additional human samples of metaplasia and cancer with nearly a 1000 patients), and in a mouse model of tumorigenesis ATF3. Together, the experiments may help us understand fundamental mechanisms cells use in regeneration and tumorigenesis that apply not just to the stomach but potentially other organs as well.